Abstract:Objective To study the impact of cSNP on the structure and function of samhd1 gene in rhesus macaques. Methods RNA was extracted from peripheral blood of rhesus macaques. The cSNP sites were detected and confirmed through reverse transcription PCR, sequencing and sequence alignment. Protein structure and function were analyzed with Predictprotein and SWISS-MODEL. Finally, the functional cSNP sites were screened by SNaPshot technique in a larger sample scale. Results Five non-synonymous mutation sites (C15G (2.17%), C320T (6.88%), G547A (13.41%), A552T (4.35%) and T839C (17.39%)) were found, respectively. Four of them, except C15G, were included in the key structure domains of SAMHD1. Meanwhile, G547A, A552T, T839C had secondary structure changed and were likely to impact the function of SAMHD1. Conclusion Some important cSNP sites are found in samhd1 gene of rhesus macaques, which may impact on the function of SAMHD1.

Abstract:Objective To investigate the PLCε gene function during growth and development of liver tumor using PLCε knockout mouse models of transplanted liver carcinoma. Methods Fifteen PLCε^+/+ mice were selected randomly as control group,and 15 PLCε^+/+(wild type)and 15 PLC^-/-(knockout genotype)as the model groups I and Ⅱ.H22 liver carcinoma cells were inoculated into the liver parenchyma of mice of the model groups by laparotomy. The mice of control group were injected with saline.Fifteen days after tumor cell inoculation, the rate of tumor formation and tumor volume were observed,and the the tumors were examined by pathology. Results The survival rate of mice in all groups was 100%.In the model group I,the tumor transplantation success rate was 100% and the mean tumor volume was (65.21±5.25)mm³.In the model group Ⅱ,the tumor transplantation success rate was 53.3% and the mean tumor volume was (23.46±3.47)mm³.The average tumor volume of the model group I was significantly greater than that of the model group Ⅱ(P<0.05).Pathological examination confirmed hepatocellular carcinoma in the model groups I and Ⅱ.Conclusion A PLCε gene knockout mouse model of transplanted liver cancer is successfully established, and it demonstrates the property that PLCε gene knockout has an inhibitory effect on liver tumor growth.

Abstract:Objective To establish a rat model of diabetic nephropathy, with similar laboratory test results and clinical symptoms in patients with diabetic nephropathy and to evaluate the stability of this animal model. Methods Sixty-six healthy male Spraque-Dawley rats were used in this study. Except those in the normal group,all rat models of diabetic nephropathy were established by unilateral nephrectomy and tail vein injection of streptozotocin (STZ).Dynamic changes of parameters such as body weight, water intake, urine volume, urinary albumin, 24 h urinary protein content, blood glucose level, glycosylated hemoglobin, blood lipid, serum creatinine and urea nitrogen were observed, and renal pathology was also examined to evaluate the success and stability of this rat models. Results The blood sugar, water intake and urine volume were higher than those in the normal group, and body weight loss was observed at 3 days to 16 weeks after STZ administration. The model group rats showed early diabetic nephropathy in 4 weeks, the urinary albumin, 24 h urinary protein and glycosylated hemoglobin were significantly increased than those in the normal group, and the blood lipid, urea nitrogen and creatinine also showed abnormality. The condition was gradually worsening with the extension of time. Conclusion Unilateral nephrectomy and tail vein injection of streptozotocin (STZ) induced nephropathy in rats is in accordance with early diabetic nephropathy in humans of both laboratory test results and clinical symptoms. The rat models at 4 to 16 weeks after modeling are in accordance with clinical nephropathy Mogensen stage Ⅲ. The severity of disease of the rat models is along with the disease course, and is stable and suitable for studies on drug pharmacodynamic evaluation and basic clinical studies on diabetic nephropathy.

Abstract:Objective To evaluate the genetic diversity of ApoE gene in Z: ZCLA gerbil closed colony. Methods PCR-SSCP technique was employed to calculate the gene frequency, genotype frequency, heterozygosity, polymorphism information content of three SNPs of ApoE gene, which were reported in our previous work, in a total of 444 ordinary and clean Mongolian gerbils. Results The average allele of 3 SNP site 97, 981 and 1774 was 2. The inheritance pattern was in accordance with Mendel's law. Expected heterozygosities were 0.063, 0.501, and 0.499, and the average expected heterozygosity of whole group was 0.354. PIC were 0.061, 0375, and 0.374, with an average PIC of 0.270. Conclusion The ApoE gene frequency and genotype distribution in Mongolian gerbils may be to a certain degree associated with the genetic drift caused by long-term closed and selected breeding.

Abstract:Objective To measure the major organ weights and body weight in Juema minipigs, calculate the organ coefficient, and analyze the correlation between body weight and major organ weights. Methods The body weight and weight of 8 organs in 21 common 7-15-month old Juema minipigs (12 females and 9 males) were measured. The organ coefficients were calculated, and sex difference was analyzed by Student's t test. Linear regression, multivariable linear regression and multiple regression analysis were carried out to analyze the correlation between body weight and major organ weights using SAS software. Results There were no significant differences between males and females in body weight, organ weights and organ coefficients. Major organ weights were positively correlated to body weight determined by correlation analysis, and the weights of heart, liver, spleen, and kidney had significant positive correlation to body weight (P<0.05), and significant linear equations were established by linear regression analysis. The weights of heart and stomach showed significant direct correlation to body weight according to the multivariable linear regression analysis, and the most close correlation was found between the heart weight and body weight. Moreover, multiple regression analysis showed that the liver, lung, kidney and bladder had significant positive correlation with heart and stomach, and spleen had significant positive correlation with liver. Conclusion There are no significant differences between major organ coefficients in male and female Juema minipigs. There exists some linear relationship between main organ weights and body weight.

Abstract:Objective To establish a quantitative method to examine the methylation level of CpG island located in the intron 1 of mouse α-synuclein, and analyze the difference of methylation in various brain regions. Methods CpG island in the intron 1 of mouse α-synuclein was identified using an online software MSPprimer. Pyrosequencing assay was designed using PyroMark assay design 2.0. DNA methylation was quantified in mouse hippocampus, cortex, striatum and midbrain using pyrosequencing assay. Results There was a CpG island in the intron 1 of mouse α-synuclein. Pyrosequencing assay measuring methylation level of this CpG island was optimized. It indicated that all of the brain regions were hypomethylated with a methylation rate of 2% or less. Conclusion DNA methylation does not play a major role in the transcription regulation of mouse α-synuclein in the brain.

Abstract:Objective To compare the impact of two anticoagulants (EDTA-3K and heparin lithium) on blood physiological parameters in miniature pigs. Methods Blood samples from 22 adult miniature pigs were collected and tested for physiological values using a MEK-7222K hematology analyzer. Differences in the values determined by using the two anticoagulants were analyzed. Results There were significant differences in values of some blood physiological parameters determined by the two anticoagulant groups, such as MON,MCV,MCHC,PLT,PCT,MPV,LYM,EOS,BAS,LYM %,MON%,EOS% and BAS%. There were significant differences in the latter 12 indexes (P<0.01) except for MON (P<0.05). Meanwhile, there were sex differences in values of 7 blood physiological indexes determined by EDTA-3K: PLT (P<0.05), NEUT (P<0.05),NEUT% (P<0.05),MON (P<0.01),EOS (P<0.01),MON% (P<0.01) and EOS% (P<0.01). There were significant differences in 8 indexes determined by heparin lithium: PDW (P<0.05), NEUT (P<0.05), LYM% (P<0.05), MON (P<0.01), EOS (P<0.01), MON% (P<0.01), NEUT% (P<0.01) and EOS% (P<0.01). Conclusion It is essential to choose proper anticoagulant before starting relevant tests, because there are significant differences in leukocyte and platelet counts in miniature pigs, determined with different anticoagulants.

Abstract:Objective To better understand the information of secreted proteins of Tupaia Chinensis.Methods The internet-based software EuSecPred 2.0 and other mapping software were collectively used to predict and analyze the genome-wide secreted proteins. Results There were 279 secreted proteins, accounting for 7.2% of the total proteins encoded by genome sequences. The function and structure of the domain annotation analysis found that they were mainly hydrolases, protein/ion binding proteins and the primary metabolic process-associated proteins. Amino acid composition analysis showed that the signal peptide secreted proteins were mainly composed of hydrophobic amino acids, and there existed some hydrophilic amino acids in the signal peptide cleavage site. Motif analysis revealed no presence of conserved motifs in signal peptide, however, five kinds of motifs existed inside the secreted proteins. Conclusion Genome-wide predictive analysis of secreted proteins of tree shrews is helpful for further study of the regulation of body functions in tree shrews, and for developing mechanism studies and animal models researches.

Abstract:Objective To study the genetic features of different populations of Kunming mice in order to enrich their genetic data and explore the method for genetic quality control. Methods SPF KM mice from two populations of Liaoning province (30 mice in each group, male:female=1:1) were used in this study. Ten microsatellite loci on different chromosomes with rich polymorphism information were chosen, and the heterozygosity (h_e), polymorphism information content (PIC) and genetic distance (D_A) were analyzed by PCR amplification and PAGE electrophoresis. Eleven morphological parameters of the mandible were determined. Results Twenty-eight alleles and 31 genotypes were found in the two mouse populations, with a h_e of 0.472 and PIC of 0.382. Twenty-seven alleles and 28 genotypes were found in the population A, with a h_e of 0.525 and PIC of 0.402. Twenty-six alleles and 27 genotypes were found in the population B, with a h_e of 0.418 and PIC of 0.361. D_A between the two populations was 0.076. The genetic diversity of group A was slightly higher than that of the group B. There were some differences in the mandibular morphology between the two populations, among them, 5 parameters had significant differences (P<0.05 or P<0.01). Conclusion The genetic distance of the two Kunming mouse populations is approximate, with a weak genetic differentiation between the two populations.

Abstract:Objective To explore the influence of carotid artery balloon injury on the formation of atherosclerotic lesions induced by hyperhomocysteinemia in rabbits. Methods Twenty New Zealand rabbits were divided into control group and model group (n=10) randomly. The left carotid arteries were injured by balloon catheterization in all rabbits. After operation, the model group was given methionine 80 mg/kg per day by subcutaneous injection for 24 weeks to induce atherosclerotic lesions, and the control group was given the same amount of normal saline. All rabbits were killed at the 24th week. ELISA was used to detect the content of homocysteine and HE staining was used to observe the pathological changes in the arterial wall. The arterial wall thickness was measured using an image analysis software. Results After 24 weeks, the level of serum homocysteine of the model group was significantly increased and pathological changes of aortic wall were observed in different degrees compared with that of the control group (P<0.05). But the carotid artery wall thickness of model group after balloon injury had no visible changes compared with that of the control group. Conclusion Balloon injury of carotid artery has no significant effect on the formation of atherosclerotic lesions induced by hyperhomocysteinemia.

Abstract:Objective To investigate if there are differences in the skin tissue structure between BALB/c mutant mice with curly hair and normal BALB/c mice. Methods 10-d,21-d,42-d and 63-d old groups of BALB/c mutant curly mice and normal BALB/c mice were used in this study, 10 mice in each group. The gross appearance of the mice were observed.Hair follicle and skin structure were examined and compared by histopathology. Results Clothing hair of the BALB/c mutant curly mice became bent at 10 day after birth,and could be distinguished from normal mice of the same age.Histological observation showed that the number of hair follicles and hair roots of BALB/c mutant curly mice was significantly less than that of normal mice, and the sparse coat and hair curling characteristics were obviously kept to 21 d, 42 d and 63 d of age.Histopathological examination revealed that the number of hair follicles in the mice in the four age groups of BALB/c mutant curly mice was also significantly less than that of normal mice.At 63 days of age, bending curly hair growing in hair follicles were visible in the BALB/c mutant curly mice. Conclusion Hair follicles and the skin tissue structure in the BALB/c mutant curly mice and normal mice showed distinct differences.These results demonstrate that the skin structure of mutant mice is also changed, which can provide a theoretical reference for future research on the mouse curly hair mutant gene and model application.

Abstract:Objective To understand some key genes and metabolic/inflammatory pathways triggered by high fat/high cholesterol (HF/HC) diet-induced hyperlipidemia in Mongolian gerbil. Methods Sixty 90-day-old Mongolian gerbils (including both sexes) were used in this study. The gerbils were divided into normal group (n=30, control diets) and model group (n=30, Hf/HC diets). Hyperlipidemia was induced for 4 weeks. To extract mRNA for transcriptome by RNA-Seq deep sequencing and bioinformatics. Results The results showed that 47 522 de novo gerbil genes (≥100 bp unigene) were obtained, with a total of 26.9 Mb, and about 82.53% sequence was homologous with the exon in the common genome bases. There were 21 125 differential genes between the hyperlipidemia gerbil and normal gerbil groups, including 16 087 down-regulated and 5038 up-regulated genes. There was a significant trend of downregulation compared with the normal group (P<0.01). There were eight pathways significantly related to the metabolic inflammation in hyperlipidemia (P<0.01), and among the expressed genes of these pathways, 15 immune-and inflammation-related genes were significantly down-regulated (P<0.01). Conclusions RNA-Seq and bioinformatics techniques can be used as a powerful tool for overall gene-transcription in hyperlipidemic gerbil models. Some key genes and pathways screened in this study may become candidate genes or operating targets in the future research.

Abstract:Objective To establish a rat model of liver fibrosis induced by bile duct ligation (BDL), and to improve the classical BDL method. Methods Eighty adult healthy male Sprague-Dawley rats were randomly divided into groups A and B, 40 rats each. The two groups of rats were modeling by classical BDL method and modified hepatic duct ligation method, respectively. Sham operation was performed in 10 rats of each group and were taken as control sham group. Hepatic biochemical indicators AST, ALT, ALP, TBIL, DBIL, GGT, and A/G in the rat serum at one week after operation were detected with an automatic biochemistry analyzer. Pathological changes of the rat liver at 4 weeks after operation were observed with HE staining. Cell proliferation-related markers α-SMA and CK-19 in the liver were detected by immunohistochemistry. Results Both two rat models of obstructive cholestasis were successfully established. Serum biochemical and liver pathological changes indicated impaired liver function in the bile duct ligated rats. After cholestasis caused by bile duct ligation, mitotic figures of hepatocytes and the expression of α-SMA and CK-19 in hepatocytes and biliary epithelial cells was increased, and hyperplasia of bile canaliculi and hepatic pseudolobule formation were observed. These changes indicated early hepatic cirrhosis. The mortality rate of rats was 66.7% in the BDL group and 26.7% in the modified BDL group. Conclusions Liver fibrosis can be successfully induced by hepatic duct ligation in rats. In addition, this modified ligation method can reduce the mortality and improve the model quality and efficacy of experiment.

Abstract:Objective To obtain higher microbiological quality of Mongolian gerbil colonies and realization of experimental animalization by biological cleaning. Methods We conducted biological cleaning of Mongolian gerbils by selection of pregnant gerbils, perinatal judgment, aseptic Caesarean section, pup milk replacer and breeding in individually ventilated cages. We also compared the results by milk replacer for pup gerbils from lactating ICR mice, rats as well as gerbils. Results During the present study, 85 Caesarean operations were carried out and a total of 167 gerbils were weaned. Conclusions We have successfully achieved the biological cleaning from a Mongolian gerbil colony in Zhejiang province, and the key techniques may provide useful reference for biological cleaning of special laboratory animal species in China.

Abstract:Due to the reduced cerebral blood supply, chronic cerebral circulation insufficiency (CCCI) causes diverse pathological damages to different degrees in the brain, and consequently leading to memory deterioration and a series of symptoms. Researches on pathological mechanism and repairing of CCCI damages are one of hot spots in neuroscience. In this article, current researches on animal models of chronic cerebral ischemia are reviewed. The authors would summarize the model preparation, evaluation of methodology and frequent questions, and to provide useful reference for further studies on chronic cerebral circulation insufficiency.

Abstract:The generation of oxidative stress is closely associated with the occurrence and development of liver injury induced by chronic alcohol consumption. It is widely considered that oxidative stress is the major mechanism of alcoholic liver disease has become a research hotspot. Nuclear factor erythroid 2-related factor 2 (Nrf2) is a key transcription factor that induces a battery of cytoprotective genes in response to oxidative/electrophilic insults. Nrf2 may play an important role in the metabolism and detoxification of a variety of drugs and chemical compounds. Deficiency of Nrf2 may aggravate the oxidative stress and damage homeostasis of redox system, resulting in cell toxicity, dysfunction, and even death. In this article we will review the research progress of Nrf2 in alcoholic liver injury models.

Abstract:As a vital component in an overall laboratory animal care and use program, development of a disaster plan plays a critical role for every research institution. Currently, most of domestic institutions would draw up an "emergency operation plan, EOP", but ignoring a practicable "business continuity plan, BCP" in establishing a disaster plan. In this article, we will discusse about the definition of disaster, how to set up an EOP, and how to establish a thorough BCP, in order to show an integrated and professional disaster plan in laboratory animal care and use program.